Tropomyosin: a new asymmetric protein component of the muscle fibril

Bailey, Kenneth

doi:10.1042/bj0430271

Cited by 631 publications

(197 citation statements)

References 18 publications

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“…For certain experiments (see Results), G-actin was further purified by gel filtration on Sephadex G-150 (41) . Tropomyosin was isolated from chicken skeletal muscle by the method of Bailey (42) as modified by Eisenberg and Kielly (43) .…”

Section: Methodsmentioning

confidence: 99%

Studies on the spectrin-like protein from the intestinal brush border, TW 260/240, and characterization of its interaction with the cytoskeleton and actin.

Pearl¹,

Fishkind²,

Mooseker³

et al. 1984

The Journal of Cell Biology

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The terminal web of the intestinal brush border contains a spectrin-like protein, TW 260/240 (Glenney, J. R ., Jr., P. Glenney, M. Osborne, and K. Weber, 1982, Cell, 28:843-854 .) that interconnects the "rootlet" ends of microvillar filament bundles in the terminal web (Hirokawa, N ., R. E. Cheng, and M . Willard, 1983, Cell, 32 :953-965; Glenney J . R ., P. Glenney, and K. , /. Cell Biol., 96:1491-1496 . We have investigated further the structural properties of TW 260/240 and the interaction of this protein with actin . Salt extraction of TW 260/240 from isolated brush borders results in a loss of terminal web cross-linkers primarily from the apical zone directly beneath the plasma membrane. Morphological studies on purified TW 260/240 using the rotary shadowing technique confirm earlier results that this protein is spectrin-like and is in the tetrameric state in buffers of low ionic strength . However, examination of TW 260/240 tetramers by negative staining revealed a molecule much straighter and more uniform in diameter than rotary-shadowed molecules . At salt concentrations at (150 mM KCI) and above (300 mM KCI) the physiological range, we observed a partial dissociation of tetramers into dimers that occurred at both 0°and 37°C. We also observed (in the presence of 75 mM KCI) a concentration-dependent self-association of TW 260/240 into sedimentable aggregates .We have studied the interaction of TW 260/240 with actin using techniques of cosedimentation, viscometry, and both light and electron microscopy. We observed that TW 260/240 can bind and cross-link actin filaments and that this interaction is salt-and pHdependent . Under optimum conditions (25-75 mM KCI, at pH 7 .0) TW 260/240 cross-linked F-actin into long, large-diameter bundles. The filaments within these bundles were tightly packed but loosely ordered . At higher pH (7.5) such bundles were not observed, although binding and cross-linking were detectable by co-sedimentation and viscometry. At higher salt (>150 mM KCI), the binding of TW 260/240 to actin was inhibited . The presence of skeletal muscle tropomyosin had no significant effect on the salt-dependent binding of TW 260/240 to F-actin .The ubiquitous presence of spectrin-like proteins in the "cortical" cytoplasm of cells has been recently established by numerous biochemical and immunological studies on a wide variety of cell types and tissues (reviewed in references 1 and 2). By analogy with erythrocyte spectrin

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Section: Methodsmentioning

confidence: 99%

Studies on the spectrin-like protein from the intestinal brush border, TW 260/240, and characterization of its interaction with the cytoskeleton and actin.

Pearl¹,

Fishkind²,

Mooseker³

et al. 1984

The Journal of Cell Biology

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“…These cDNAs have been previously described (Goodwin et al, 1991). The TM proteins were purified from the bacterial lysates using modifications to those methods described for isolation of tropomyosin from tissue (Bailey, 1948;Bourdillon and Baker, 1956; see Materials and Methods). The proteins were analyzed biochemically by SDS-PAGE, amino acid analysis, amino-and carboxy-terminal sequence analysis, and their ability to bind to F-actin.…”

Section: Preparation and Characterization Of Bacterially Produced Tmsmentioning

confidence: 99%

In vitro and in vivo characterization of four fibroblast tropomyosins produced in bacteria: TM-2, TM-3, TM-5a, and TM-5b are co-localized in interphase fibroblasts.

Pittenger

Helfman

1992

The Journal of Cell Biology

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Abstract. Most cell types express several tropomyosin isoforms, the individual functions of which are poorly understood. In rat fibroblasts there are at least six isoforms; TM-1, TM-2, TM-3, TM-4, TM-5a, and TM-5b. TM-1 is the product of the/5 gene. TM-4 is produced from the TM-4 gene, and TMs 2, 3, 5a, and 5b are the products of the oe gene. To begin to study the localization and function of the isoforms in fibroblasts, cDNAs for TM isoforms 2, 3, 5a, and 5b were placed into bacterial expression vectors and used to produce TM isoforms. The bacterially produced TMs were determined to be full length by sequencing the amino-and carboxy termini. These TMs were found to bind to F-actin in vitro, with properties similar to that of skeletal muscle TM. In addition, competition experiments demonstrated that TM-5b was better than TM-5a in displacing other TM isoforms from F-actin in vitro. To investigate the intracellular localization of these fibroblast isoforms, each was derivatized with a fluorescent chromophore and microinjected into rat fibroblasts. TM-2, TM-3, TM-5a, and TM-5b were each found to associate along actin filaments. There was no preferred cellular location or subset of actin filaments for these isoforms. Furthermore, coinjection of two isoforms labeled with different fluorochromes showed identical staining. At the level of the light microscope, these isoforms from the ct gene do not appear to achieve different functions by binding to particular subsets of actin filaments or locations in cells. Some alternative possibilities are discussed. The results show that bacterially produced TMs can be used to study in vitro and in vivo properties of the isoforms.

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“…Ebashi and Kodama (l965) subsequently found that "native tropomyosin" actually consisted of two proteins. One of these two proteins was tropomyosin, which had been known to be a component of the myofibril since 1948 when it was discovered by Bailey (Bailey, 1948). The physiological function of tropomyosin, however, had remained unclear ever since its discovery.…”

Section: +mentioning

confidence: 99%

Localization of [alpha]-actinin and tropomyosin in muscle and nonmuscle systems

Schollmeyer¹

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Tropomyosin: a new asymmetric protein component of the muscle fibril

Cited by 631 publications

References 18 publications

Studies on the spectrin-like protein from the intestinal brush border, TW 260/240, and characterization of its interaction with the cytoskeleton and actin.

Studies on the spectrin-like protein from the intestinal brush border, TW 260/240, and characterization of its interaction with the cytoskeleton and actin.

In vitro and in vivo characterization of four fibroblast tropomyosins produced in bacteria: TM-2, TM-3, TM-5a, and TM-5b are co-localized in interphase fibroblasts.

Localization of [alpha]-actinin and tropomyosin in muscle and nonmuscle systems

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